Project Summary Diffuse intrinsic pontine glioma (DIPG) the most common pediatric brainstem tumor and is almost uniformly fatal. This disease has a devastating 9 month median survival after diagnosis, despite over 200 clinical trials. Based upon several lines of clinical and preclinical evidence, we believe that delivering intratumoral Reolysin, an oncolytic virus, can improve survival outcomes in patients with DIPG. Oncolytic viruses include a variety of natural and engineered viruses that elicit varying combinations of direct tumor lysis and local/systemic antitumor immunity. This proposal will use Reolysin, a wild type Dearing type 3 reovirus developed to clinical grade by Oncolytics Biotech. Preclinical studies and Phase 1 clinical trials demonstrated intracranial Reolysin's safety and antitumor activity. Our preliminary studies for this grant also show that Reolysin can replicate and lyse DIPG cells in vitro. However, DIPG's sensitive location in the brainstem creates the potential for inflammatory toxicity. Additionally, to translate this this work to a clinical venue and also address NCI's Provocative Question #11 (assessing the impact of clinical standard of care on immunotherapy), we need to verify this therapy works in the clinical standard of care setting for DIPG, including local radiotherapy (RT) and systemic dexamethasone (DEX). Reolysin therapy may be potentiated or altered by either RT, which causes cell death, or DEX, which is an immunomodulatory agent. The first half of the project will measure the direct oncolysis, local/systemic antitumor immunity, and efficacy/toxicity of direct Reovirus administration both in immunodeficient and immunocompetent murine models. The second half of the project will use a combination of in vitro studies and data generated in the first half of the project to incorporate the clinical standard of care (RT/DEX) and develop a treatment regimen that maximizes efficacy and limits toxicity by varying dosages and timing of administration. If a viable regimen is created, it will be used to guide and direct future clinical trials, and if not, there are many other oncolytic viruses with varying therapeutic profiles that could be studied using the same models and techniques.